| Objective： To select carbapenem-resistant Acinetobacter baumannii(CRAB), research the role of carbapenemases and efflux pump genes inCRAB from gene level, analyze the homology of CRAB strains inChengdu region and to provide important evidence for nosocomialinfection control in the region.Methods:1.160clinical and environmental isolates of Acinetobacterbaumannii complex were collected. All of isolates were rapidly identifiedby a multiplex polymerase chain reaction(PCR).2. All Acinetobacterbaumannii isolates underwent antimicrobial susceptibility testing of14antibiotics for selecting CRAB isolates.3. The CRAB isolates wereexamined for the presence of oxacillinases(OXA-23, OXA-24, OXA-51,OXA-58and OXA-143) by a multiplex PCR， andmetallo-β-lactamases(NDM-1, IMP, VIM and SIM) were detected by PCR.Insertion sequence ISAba1and the relationship with OXA were detected byPCR.5. Efflux pump AdeABC, AdeIJK and AbeM were detected by PCR.The expression of efflux pump AdeABC, AdeIJK and AbeM of carbapenem-sensitive and resistant Acinetobacter baumannii isolates wereanalyzed by real-time PCR.6. Homology analysis of CRAB strains wasperformed by DiversiLab system.Results：1.The result showed that132(132/160,82.5%) isolates wereidentified to Acinetobacter baumannii by a mutiplex PCR2.79out of132isolates were CRAB strains. The rate of resistance was59.84%. The resultsof antimicrobial susceptibility testing suggested that there was a highresistance rate to common used antibiotics. Cefoperazone/sulbactam andcefepime resistance rate were79.9%,84.8%respectively. The rest ofantibiotics resistance rate was more than90%.3. All CRAB strainspossessed OXA-51gene and OXA-23gene, and only OXA-51wasdetected in CSAB isolates, whereas no isolates carried OXA-24, OXA-58，OXA-143or the metallo-β-lactamases except one isolates detected IMP. Allthose indicated that OXA-23was one of mechanisms in CRAB isolates.ISAba1was identified in79CRAB isolates. The structure ISAba1-OXA-23was detected in77CRAB isolates. ISAba1lay upstream of OXA-23in77CRAB isolates by sequencing and blasting. We also found that the77CRAB strains containing ISAba1-OXA-23structure was completelyresistant to carbapenems, while2CRAB strains without ISAba1-OXA-23were sensitive to one of the carbapenem antibiotics. All suggested thatISAba1-OXA-23structure might be associated with the CRAB resistance.4. The positive rates of amplified the efflux pump AdeB, AdeJ and AbeM in79CRAB isolates were58.2%,96.2%and97.5%respectively, while thepositive rates of amplified the efflux pump AdeB, AdeJ and AbeM in20CSAB isolates were25%,85%and95%respectively. All suggested that itmight present the efflux pump gene between CRAB with CSAB strains.However, the expression of efflux pump genes and regulation of them maybe related to CRAB.5. Real-time PCR identified5.59-fold increases inAdeJ expression in CRAB compared to CSAB. However, there were nodifference in AdeB and AbeM expression between CRAB and CSAB. Itsuggested that the efflux pump AdeJ might be related to CRAB resistance.6. The representative40isolates were separated into four unrelated groupsand nine patterns by DiversiLab system, with the highest prevalence ofgroup1being found in five hospitals. It suggested that the popular CRABclone1might be disseminated throughout the region.Conclusion:1. Acinetobacter baumannii could be rapidly and accuratelyidentified by multiplex PCR.2. The Acinetobacter baumannii isolates werewidely resistant to carbapenem and the phenomenon of multi-drug resistantA.baumannii was serious.3. Production OXA-23was one of importantmechanisms of Acinetobacter baumannii resistance to carbapenem. ISAba1lay upstream of OXA-23and might be involved in the regulation of theexpression of OXA-23.4. Efflux pump genes were be detected in CRABand CSAB isolates. Overexpression of AdeIJK might be one of reasons ofCRAB.5. The same clone group of CRAB presented within or between hospitals in Chengdu area. We should take some measures to controlAcinetobacter baumannii infection throughout the whole region. |
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